Mutations in the clotting factor VIII (fVIII) gene result in a decreased or defective clotting factor (fVIII) protein that gives rise to hemophilia A, which is characterized by uncontrolled bleeding. Hemophilia B is similarly associated with clotting factor IX (fix). Treatment of hemophilia A typically entails lifelong, multi-weekly intravenous infusion of either human plasma-derived or recombinant fVIII product. Due to the high cost, less than 30% of the global hemophilia A population receives this form of treatment. Furthermore, about 25% of patients treated with fVIII replacement products develop neutralizing antibodies that render future treatment ineffective. Thus, there is a need to identify improved therapies.
Gene therapies are typically based on genetically engineering viruses designed to deliver functional transgenes to the patient so that their own cells can biosynthesize missing or defective proteins. Clinical advancements have been made using recombinant adeno-associated viral (rAAV) vectors for the expression of fIX in the liver; however, using rAAV for fVIII expression for patients with hemophilia A has been challenging due to inefficient biosynthesis of human fVIII (hfVIII). Recombinant adeno-associated viral (rAAV) vectors produce capsids that have a limited space for encapsulating nucleic acids. FVIII is a large glycoprotein, and the rAAV sequences needed for encoding and expressing fVIII typically exceed capsid packing capacity.